Eukaryotic cells are organized with separate membrane-bound organelles with distinct lipid compositions. The lipid compositions are maintained through vesicular transport as well as the less-understood, lipid-transfer protein (LTP)-mediated, nonvesicular transport. It is important to gain a better understanding of lipid transfer process, as perturbations of lipid trafficking contribute to human pathologies including cancer, neurodegenerative disorders, cardiovascular diseases, obesity and diabetes. Viruses, as obligate intracellular parasites, have evolved strategies to manipulate the cellular membranes for entry, genome replication, virion production, and exit. Uncovering these strategies will not only reveal key viral replication steps for antivirals development but also provide mechanistic insights on fundamental cellular processes. Enveloped viruses typically acquire their outer lipid bilayer by budding from cellular membranes, a process that is similar to the formation of cellular transport vesicles. Poxviruses, however, are unusual in that their primary envelope is not acquired by budding but through extending of open-ended crescent membranes. The origin and biogenesis of the crescent membranes have puzzled virologists for over half a century, albeit recent studies suggest that the crescents may derive from the endoplasmic reticulum (ER) in a manner that is independent of vesicular transport from the ER. Five viral proteins, collectively termed viral membrane assembly proteins (VMAPs), have been found to be essential for the biogenesis of crescent membranes. The A6 protein of vaccinia virus is a key member of the VMAPs, which we discovered and have studied intensively. In recent studies, we achieved a breakthrough in structural analysis of the A6 protein by solving the crystal structures of both its N- and C- domains. Even more importantly, our structural and biochemical studies indicate that the C- domain is a novel lipid binding protein with an unusually high binding capacity for glycerol-phospholipids and that the N-domain regulates lipid binding. These led us to the innovative hypotheses that A6 is a lipid-transfer protein (LTP) and that poxviruses obtain their primary envelope by mimicking or hijacking the cellular LTP-mediated nonvesicular lipid transport process. We propose the following exploratory studies to test our novel hypotheses.
Aim 1. To determine the specificity and stoichiometry of the lipids bound by A6 Aim 2. To determine the role of lipid binding of A6 in viral membrane biogenesis Aim 3. To determine the structural basis by which A6 N-domain regulates C-domain for lipid binding.
Pathogenic poxviruses such as variola virus and monkey pox virus remain a dangerous threat to today's largely unimmunized populations. This project of studying poxvirus membrane biogenesis will reveal key viral replication steps for antivirals development and provide fundamental knowledge on cellular lipid trafficking processes. Perturbations of lipid trafficking contribute to human pathologies including cancer, neurodegenerative disorders, cardiovascular diseases, obesity and diabetes.
| Pathak, Prabhat Kumar; Peng, Shuxia; Meng, Xiangzhi et al. (2018) Structure of a lipid-bound viral membrane assembly protein reveals a modality for enclosing the lipid bilayer. Proc Natl Acad Sci U S A 115:7028-7032 |
| Khandelwal, Anuj; Kent, Caitlin N; Balch, Maurie et al. (2018) Structure-guided design of an Hsp90? N-terminal isoform-selective inhibitor. Nat Commun 9:425 |
